The goal of this project is to better understand the effects of host cells and other co-factors on human retroviral replication and pathogenesis. Highly active combination antiviral drug therapy (HAART) holds promise to convert productive HIV infections into a chronic, non-viremic, non-progressive condition. However, in some patients, limited but continuous viral production has been observed and, in others, latently infected long-lived peripheral T cells which can be reactivated during a normal immune response have been identified in HAART-treated HIV infected individuals. In developing long term therapy for HIV infected individuals it will be necessary to control viral production from and/or eradicate these latent reservoirs through adjuvant therapies like immunotherapy. We have previously identified two mechanisms which activate expression of latent HIV provirus: immune activation, which requires contact between latently infected cells and uninfected activated CD4+ cells, and hypomethylation. We have recently demonstrated that acute HIV infection increases the cellular capacity to methylate genes. Increased DNA methyltransferase (DNMT) activity in HIV infected cells can result in an increase in the latent reservoir through methylation of the HIV LTR and the subsequent silencing of viral gene expression. In addition, increased DNMT activity can result in aberrant methylation of cellular genes including, interferon-gamma (IFN-gamma). This hypermethylation of IFN-gamma results in decreased production of IFN-gamma, decreased type 1 immune response and increased spread of cell free virions. In attempting to understand the role of methylation in the regulation of viral and gene regulation, we have made progress in several areas. First, HIV proviral integration is not necessary for HIV infection to increase the cellular capacity to methylate genes. Second, DNMT-1 but not DNMT3a and DNMT3b are increased during HIV infection of lymphoid cells. Third, in some (49/96) patients on long-term HAART, CD4+ T cells have greatly diminished ability to produce IFN-gamma which can be restored through hypomethylation of IFN-gamma promoter,using hypomethylating drugs. Bisulfite genomic sequencing of this promoter has shown that the IFN-gamma promoter can remain hypermethylated during HAART therapy. Fourth, using a newly developed array based assay (differential methylation hybridization), we have found that CD4+ T lymphocytes from HIV infected individuals have several CpG islands which are more extensively methylated than normal controls. We have identified one of these genes as the p16(Ink4a) geneindicating that CpG islands, normally hypomethylated, can be hypermethlated after HIV infection. Since DNMT1 is the DNMT that is predominantly upregulated following HIV infection and recent evidence suggests that, in addition to its enzymatic function, it can form complexes with otherproteins including transcriptional factors such as Tat or Vpr can tether to complexes containing DNMT-1 dysregulate transcription control in a novel manner. Although largely unknown, the cellular events triggered by HTLV-1 infection in the lymphoproliferative process are of major importance to the establishment of leukemia. We have recently characterized an infectious clone of HTLV-1 as well as a susceptible expressing cell line to study the molecular interactions of this infectious clone. The HTLV-1 surface unit of the envelope has been fused to the Ig molecule and is being used to study the regulation of the HIV-1 receptor. This project was formerly Z01 CM 09251-12 LLB. AIDS Title: Negative Regulation of HIV replication - Pathogenic Effects